Chapter 3 Protein Structure & Function

Question 1

What are the 3 most widely used characteristics for separating proteins?

Answer 1

1) size (length or mass)
2) net electrical charge
3) affinity for specific ligands

Question 2

What is the first step in a typical protein purification scheme?

Answer 2

Centrifugation

Question 3

What is the principle behind centrifugation?

Answer 3

Two types of particles in suspension will with different masses or densities will settle to the bottom of a test tube at different rates

Question 4

Proteins vary greatly in _____ but not in _____

Answer 4

1) Mass

2) Density

Question 5

Two basic purposes of centrifugation

Answer 5

1) preparative technique to obtain enough of material for further experiments
2) analytical technique to measure physical properties of macromolecules

Question 6

What is the most common initial step in protein purification from cells or tissues?

Answer 6

Differential centrifugation - the separation of water soluble proteins in insoluble cellular material

Question 7

What is cell homogenate?

Answer 7

Mechanically broken cells. It is the starting mixture used in differential centrifugation.

Question 8

What is the primary structure of protein?

Answer 8

The linear arrangement or sequence of amino acids

Question 9

What is the secondary structure of protein? What type of bonds?

Answer 9

• The various spatial arrangements that result from folding localized regions of the polypeptide chain
• include the alpha helix and the beta sheet
• are held together by hydrogen bonds

Question 10

What is the tertiary structure of protein? What type of bonds?

Answer 10

• the overall conformation of the polypeptide chain, its three-dimensional structure
• primarily stabilized by hydrophobic interactions between non-polar side chains of the amino acids and hydrogen bonds between polar side chains

Question 11

What is the quaternary structure of protein

Answer 11

the number and relative positions of the subunits in a multimeric protein

Question 12

4 features of mass spectrometer

Answer 12

1) the ion source
2) the mass analyzer
3) the detector
4) a computerized data system

Question 13

What is X-ray crystallography? How does it work?

Answer 13

• used to determine the 3D structure of proteins
• x-rays are passed through a protein crystal
• the diffraction pattern generated when atoms in the protein scatter the x-rays is a characteristic pattern that can be interpreted into defined structures

Question 14

What is Cryoelectron microscopy? How does it work?

Answer 14

• rapid freezing of a protein sample and examination
  with a cryoelectron microscope
• A low dose of electrons is used to generate a scatter pattern that can be used to reconstruct the protein’s structure

Question 15

What is nuclear magnetic resonance (NMR) spectroscopy? How does it work?

Answer 15

a protein solution is placed in a magnetic field and the effects of different radio frequencies on the spin of
different atoms are measured. From the magnitude of the effect of one atom on an adjacent atom, the distances between residues can be calculated to generate a 3D structure.

Question 16

Advantages and disadvantages of x-ray chrystallography?

Answer 16

(Advantage) can provide extremely high-resolution structural information on molecules and molecular complexes of any size. So long as a suitable crystal can be obtained, this is ideal for large proteins and macromolecular assemblies.

(Disadvantage) the challenge of producing samples in the form of single crystals suitable for diffraction experiments.

Question 17

Advantages and disadvantages of cryoelectron microscopy?

Answer 17

(Advantage) the relative ease of sample preparation. As long as a suitable crystal can be obtained, this is ideal
for large proteins and macromolecular assemblies.

(Disadvantage) structural resolution is generally not so high as with the other methods, especially for asymmetric assemblies.

Question 18

Advantagesand disadvantages of NMR spectroscopy?

Answer 18

(Advantage) gives high-resolution information on protein structures in solution. better for small proteins.
Ideal for monitoring protein dynamics

(Disadvantage) limited in its ability to conclusively determine the structures of very large proteins and symmetrical macromolecular assemblies.

Question 19

Does the addition of an enzyme affect the free energy of the substrate or product?

Answer 19

No. Therefore the difference in free energy for a chemical does not change as a result of adding an enzyme

Question 20

What is ubiquitin?

Answer 20

A 76-amino acid protein that serves as a molecular tag for proteins destined for degradation.

Question 21

What is ubiquitination?

Answer 21

Ubiquitination of a protein involves an enzymecatalyzed
transfer of a single ubiquitin molecule to the lysine side chain of a target protein. This ubiquitination step is repeated many times, resulting in a long chain of ubiquitin molecules. The resulting polyubiquitin chain is recognized by the proteasome, which is a large, cylindrical, multisubunit complex that proteolytically
cleaves ubiquitin-tagged proteins into short peptides and free ubiquitin molecules.

Question 22

What is Cooperativity?

Answer 22

• aka. allostery
• Any change in the tertiary or quaternary structure of a protein induced by the binding of a ligand that affects the binding of subsequent ligand molecules a multisubunit protein can respond more efficiently to small changes in ligand concentration compared to a protein that does not show cooperativity

Question 23

What catalyzes phosphorylation (the addition of phosphate groups) ?

Answer 23

protein kinases

Question 24

What catalyzes dephosphorylation (removal of phosphate groups) ?

Answer 24

protein phosphatases

Question 25

active site

Answer 25

Specific region of an enzyme that binds a substrate

molecule(s) and promotes a chemical change in the bound substrate. (Figure 3-23)

Question 26

alpha helix

Answer 26

Common protein secondary structure in which the linear sequence of amino acids is folded into a right-handed spiral stabilized by hydrogen bonds between carboxyl and amide groups in the backbone. (Figure 3-4)

Question 27

autoradiography

Answer 27

Technique for visualizing radioactive molecules
in a sample (e.g., a tissue section or electrophoretic gel) by exposing a photographic film (emulsion) or two-dimensional electronic detector to the sample. The exposed film is called an autoradiogram or autoradiograph.

Question 28

beta sheet

Answer 28

A flat secondary structure in proteins that is created by hydrogen bonding between the backbone atoms in two different polypeptide chains or segments of a single folded chain.(Figure 3-5)

Question 29

beta turn

Answer 29

A short U-shaped secondary structure in proteins.

Figure 3-6

Question 30

chaperone

Answer 30

Collective term for two types of proteins—molecular
chaperones and chaperonins—that prevent misfolding of a target protein or actively facilitate proper folding of an incompletely folded target protein, respectively. (Figures 3-17 and 3-18)

Question 31

conformation

Answer 31

The precise shape of a protein or other macromolecule
in three dimensions resulting from the spatial location
of the atoms in the molecule. (Figure 3-8)

Question 32

domain

Answer 32

A region of protein that has a distinct, and often independent, function or structure, or that has a distinct topology relative to the rest of the protein.

Question 33

electrophoresis

Answer 33

Any of several techniques for separating macromolecules based on their migration in a gel or other medium subjected to a strong electric field. (Figure 3-38)

Question 34

enzyme

Answer 34

A protein that catalyzes a particular chemical reaction

involving a specific substrate or small number of related substrates.

Question 35

homology

Answer 35

Similarity in characteristics (e.g., protein and nucleic
acid sequences or the structure of an organ) that reflects a common evolutionary origin. Proteins or genes that exhibit homology are said to be homologous and sometimes are called homologs. In contrast, analogy is a similarity in structure or function that does not reflect a common evolutionary origin.

Question 36

kinase

Answer 36

An enzyme that transfers the terminal (g) phosphate
group from ATP to a substrate. Protein kinases, which phosphorylate specific serine, threonine, or tyrosine residues, play a critical role in regulating the activity of many cellular proteins. See also phosphatases. (Figure 3-33)

Question 37

ligand

Answer 37

Any molecule, other than an enzyme substrate, that binds tightly and specifically to a macromolecule, usually a protein, forming a macromolecule-ligand complex.

Question 38

peptide bond

Answer 38

The covalent amide linkage between amino acids
formed between the amino group of one amino acid and the carboxyl group of another with the net release of a water molecule (dehydration). (Figure 2-13)

Question 39

phosphorylation

Answer 39

The covalent addition of a phosphate group to
a molecule such as a sugar or a protein. The hydrolysis of ATP often accompanies phosphorylation, providing energy to drive the reaction and the phosphate group that is covalently added to the target molecule. Enzymes that catalyze phosphorylation are called kinases.

Question 40

polypeptide

Answer 40

Linear polymer of amino acids connected by peptide

bonds, usually containing 20 or more residues.

Question 41

proteasome

Answer 41

Large multifunctional protease complex in the cytosol that degrades intracellular proteins marked for destruction
by attachment of multiple ubiquitin molecules. (Figure 3-31)

Question 42

protein

Answer 42

A macromolecule composed of one or more linear polypeptide chains and folded into a characteristic three-dimensional shape (conformation) in its native, biologically active state

Question 43

proteome

Answer 43

All the proteins in a cellular compartment, intact cell,

organ, or organism.

Question 44

proteomics

Answer 44

The systematic study of the amounts, modifications,
interactions, localization, and functions of all or subsets of proteins at the whole-organism, tissue, cellular, and subcellular levels

Question 45

structural motif

Answer 45

A particular combination of two or more secondary
structures that form a distinct three-dimensional structure that appears in multiple proteins and that often, but not always, is associated with a specific function.